Tardigrades, also known as “water bears” or “moss piglets,” are microscopic animals that have captured the public’s imagination in such forums as the new Star Trek: Discovery television series (https://www.space.com/38477-star-trek-discovery-mudd-ies-up-tardigrade-science.html) for their ability to survive the radiation of space (Rebecchi et al. 2009). Although most microbiologists would consider them microfauna, or microscopic animal life, rather than microbes strictly speaking, we are including them here because they are invisible to the naked eye. You typically need a microscope to see these creatures less than one millimeter long.However, like humans rather than bacteria, tardigrades have nuclei in their cells, and are even multicellular – that is, they are made up of many cells (Nichols 2005). Tardigrades have a piercing mouth part that some herbivorous species use to eat mosses or plant-like algae, and other predatory species use to eat microscopic animals such as nematodes or rotifers (Nichols 2005, Hohberg & Traunspurger 2009).

When placed on a glass or plastic petri dish or slide to view under a microscope, tardigrades have a hard time gaining traction, and thus look like they are running in place, moving slowly: hence the name tardigrade, which means “slow stepper.” However, if they have sand or something sticky in a petri dish like agar, they can walk somewhat like an eight-legged caterpillar might. They live in many environments around the world, especially in mosses, in leaf litter, in the nutrient-poor soils of polar and alpine environments, and can even be found in the ocean (Nichols 2005).​The tardigrades pictured here were extracted from cryoconite holes, or small mud puddles, on Antarctic glaciers.

Phialophora is a genus of fungus containing taxa capable of a variety of functions. They can be parasitic on fruits and humans, saprotrophic (decompose dead plant material), or mutualistic with plants. At our field site at Niwot Ridge in the Colorado Rocky Mountains, Phialophora was one of the most abundant fungi we found living inside of plant roots. We found DNA sequences of this genus in 126 of 177 plants that we sampled. Before extracting the DNA from plant roots, we washed them in ethanol and bleach to sterilize the root surface; thus we believe this fungus is inhabiting the inside of plant roots and directly interacting with the plant.

Fig. 1. Phialophora gregata inside of a Ranunculus adoneus root. The brown color is typical of dark septate fungi. The storage structures seen here are called microsclerotia. From Schadt et al. 2001.

We are interested in this fungus because it is a dark septate fungus that can benefit plant growth. For example, Newsham (1999) found that the fungus Phialophora graminicola benefited the growth of its host plant. We’ve known that this group of fungi lives in the alpine for a while now. A former student in the lab, Chris Schadt, found a fungus in this genus (Phialophora gregata) inside of an alpine plant back in 2001 (Fig. 1). Interestingly, the sequences from our 2016 survey at Niwot were closely related to Phialophora that were found inside of moss tissues on King George Island in Antarctica, which suggests that this genus is widespread geographically. We are currently trying to culture this fungus to isolate it and conduct experiments with it in the greenhouse to see if this particular strain benefits plant growth.

Mortierella species are aseptate, filamentous soil fungi belonging to the order of Mortierellales of the Zygomycota phylum. Among the 70+ described species, most are commonly found in soil. As saprophytic organisms (those that live on dead or decomposing matter), they can be associated with a multitude of substrate sources including plant litter. A unique feature of Mortierella is that some species are known to be exceptionally well adapted to living under very cold conditions typical of arctic, Antarctic and high alpine snow-covered soils. As psychrophilic (cold-loving), they can grow at near- or even below-freezing temperatures and survive freeze-thaws cycles and desiccation. To tolerate cold, they employ physiological mechanisms that diminish the possibility of cellular water to freeze, crystalize, and rupture cell membranes. The most common mechanisms include production of trehalose and cryoprotectant sugars that stabilize membranes, glycerol and mannitol that maintain turgor pressure, unsaturated fatty acids that maintain fluidity of membrane structures, and antifreeze proteins that may slow the growth of ice crystals.

Fig. 1. A Mortierella fungus growing on a plate in the lab.

At our field site at Niwot Ridge in the Colorado Rocky Mountains, Mortierella species can be easily observed as white mats, also known as “snow molds”, that quickly appear at the edges of receding snow banks in the spring time. Their incredible fast growth rates have been attributed to their ability to rapidly exploit a fresh flush of nutrients stored in soil and water from melting snow. However, as soon snow is gone, they disappear as fast as they appeared. In a recent survey of soil communities from a range of soil habitats spanning bare to increasingly vegetated, the abundance of Mortierella spp. declined where plant communities have become more established. Because increasing incidence of plants in this landscape reflects diminishing snow cover likely due to climate warming, the future of cold-loving Mortierella spp. lies in “hands” of surviving snow packs keeping high alpine ecosystems free of vegetation.

The microbe we will be covering this month comes to us from our nearest neighboring domain on the tree of life: The Archaea. While many think of the Archaea rare in today’s world, the truth is that Archaeal species can be found in almost every environment we look. Nitrososphaera viennensis can even be found in everyday soils that are located in our own backyards, and was originally isolated from garden soil in Vienna, Austria (Stieglmeier et al. 2014).

While displaying unique morphological traits (Figure 1) N. viennensis is interesting to researches and the public alike for its ability to feed on ammonium (NH4+). Research has already shown that a close relative of N. viennensis known as Nitrososphaera gargensis can be a potential tool in combating nitrogen pollution in water[1]. N. viennensis has even been found in manure piles produced from farmland waste[5] and may be important in the breakdown of ammonium in fertilizers. I would like to highlight that due to the worldwide distribution of these organisms, and their relatives, they play a key role in the global nitrogen cycle. Nitrogen pollution produced by human activity has severally altered the world’s nitrogen cycle[2]. Thus understanding how these organisms’ ecology will help us understand how we have impacted the natural world and how we might be able to look back to that world to find solutions.

Every living thing needs it, but nitrogen (N) is tough to get in nature. On earth our primary source of N is the air, which is 80% N. But atmospheric N is very difficult for living things to use, because it consists of two N atoms triple-bonded together. Getting them apart, so they can be bonded to other atoms, requires a LOT of energy. Fortunately, Cyanobacteria of the genus Nostoc are famous worldwide for their ability to convert atmospheric N into a biologically useful form. Although other microbes can do this “N-fixing” too, these photosynthetic bacteria are special because they fix N, they fix C (they turn atmospheric carbon dioxide into their food!), and they are really cool-looking.

Some of these Nostoc grow as spherical colonies (see picture), which are eaten by people in Peru and Chile. We call these strains Nostoc commune var. sphaeroides, but in Peru they are called “algas de la laguna”, “cushuro”, and “llullucho”. The edible forms of these cyanobacteria are commonly found in streams, but their close relatives have been found in much more extreme environments. In our expeditions to the Peruvian Andes, we’ve found tiny (less than 1mm in diameter) Nostoc spheres in dry, plant-free soil that was recently uncovered by a retreating glacier (Darcy et al. 2018).

We also found related Nostoc species at extremely high-elevation, above 5500 meters above sea level (18,100 feet), where the air is very cold, it’s very dry, and only the toughest microbes can survive (Schmidt et al. 2017).

September’s microbe of the month is the genus of fungi Aspergillus. Aspergillus is a mold type of fungus and is another fungus that we’ve found in the roots of alpine plants on Niwot Ridge, Colorado. However, the genus is very widespread, and can be found indoors as well.

Interestingly, it was also one of the few fungal taxa that we found in air samples that we collected at Niwot Ridge last summer. We put up air filters (Figure 1) that trap fungal spores and other microbes to examine potential dispersal limitation of microbes. In other words, are microbes able to be blown all over the landscape, or are they only able to reach some random locations?

Figure 1. Passive air filter for sampling airborne microbes.

​Unlike some of the other fungi I have written about, Aspergillus is typically pathogenic to plants and can even cause disease in humans and other animals too. While mycorrhizae take carbon from plants, they give the plants nutrients in return and have a beneficial effect. Pathogens like Aspergillus take resources from plants without giving them anything in return. One of the most abundant species in the genus, Aspergillus fumigatus, can cause pulmonary infections in humans. There has been a lot of work on Aspergillus in the agricultural field, as some species have been known to cause disease on important crops such as corn.

In keeping with the summer’s fungus theme, this month’s microbe is Amanita muscaria, another one that we see commonly in the Colorado subalpine on our way up to sites at the Niwot Ridge Long Term Ecological Research site. Amanita is straight out of the Super Mario games or Alice in Wonderland – the classic red mushroom with white spots. Unlike the King Bolete I wrote about last month, I do not recommend eating the Amanita, as it is highly toxic, containing ibotenic acid and muscimol compounds. In contrast to the spongy King Bolete, the Amanita has gills on the underside of the cap, which is the first sign that you should be wary of eating it.

Amanita muscaria is one of 600 species in the Amanita genus. They are in the Amanitaceae family, in the Agaricales order (gilled mushrooms) in the Basidiomycota phylum. Amanita muscaria is mycorrhizal, and associates with both hardwoods and conifers. In Colorado, it appears to be associating with the subalpine fir, all the way up to treeline, and produces mushrooms in July and August after it rains. It ranges from Mexico all the way to Alaska. The cap is 5-25 cm in diameter, deep to bright red, with yellow “warts” that quickly fad to white.

Hello! This month’s microbe is a tasty one – the King Bolete (or Porcini) mushroom (Boletus edulis). At our Colorado field site, we have been seeing these mushrooms pop up in the subalpine as we walk up to our plots! It is abundant in the Colorado subalpine forests because of its association with spruce trees. The fungus forms a symbiotic ectomycorrhizal association, meaning it envelops roots but does not penetrate them extensively like an arbuscular mycorrhizal fungus. B. edulis, like other ectomycorrhizae, helps trees acquire nutrients and can also increase the ability of seedlings to resist water stress.

B. edulis is basidiomycete fungus (Phylum Basidiomycota) that is widely distributed across Europe, Asia, and North America, where it associates with a variety of tree species besides spruce. Fruiting bodies (mushrooms) of the fungus appear in the summer to fall, especially after rain and wet conditions. Drought, low humidity, and low night air temperatures and frost events inhibit the appearance of the fruit bodies. The cap of the mushroom ranges from 7 to 30 cm in diameter. The underside of the cap has a spongey, not gilly, texture. The mushrooms are edible and are widely renowned for their great taste.

Hello! This month’s microbe the genus Entrophospora. Entrophospora is an arbuscular mycorrhizal fungus (AMF). What does that mean? A mycorrhizal fungus is any fungus that grows on or inside of plant roots and directly interacts with the plant in some capacity, typically nutrient exchange. There are several different types of mycorrhizae such as:

Ectomycorrhizae- which live on the outside of plant roots and are common associates of many tree species. Ericoid mycorrhizae- which associate with plants in the Ericaceae family such as blueberries and cranberries. Dark septate endophytes- which are described in detail in the April 2017 post below.

Arbuscular mycorrhizae live on the inside of plant roots and their defining feature is the formation of treelike structures called arbuscules (Figure 1). Arbuscules are the site of active two-way transfers of nutrients between the plant host and the fungus(1). All arbuscular mycorrhizae are in the phylum Glomeromycota(2), which is a separate phylum from typical edible mushrooms you may eat (typically Ascomycota or Basidiomycota).

Figure 1. AMF structures inside a plant root. Panel A shows blue stained hyphae invading the brick-like plant root, and 2 arbuscules. Panel B is a blown-up image of an arbuscule. Panel A taken from https://mycorrhizas.info/vam.html; Panel B taken from http://www.gpnmag.com/article/mycorrhizae-description-of-types-benefits-and-uses/ .

Entrophospora was the most abundant of the 28 AMF genera that we found in our 2016 Colorado root samples, based on DNA analysis. We found DNA matching Entrophospora in 27 plant samples. Entrophospora, like other AMF at our sites, is likely helping plants acquire phosphorus. Previous work in our lab showed correlations between arbuscules and plant phosphorus levels1, which supports this suggestion. Undeveloped soil, typical in high elevation sites recently exposed by receding glaciers, is often very limited in phosphorus, as it is still locked up in bedrock and has not had time to become available(3).

This month’s microbe, Paramecium putrinum, is a ciliate. Ciliates are single celled animals, but can be large by microbial standards: many of them that I have observed are more than 100 µm long. That may sound small, but it is 50 times larger than most bacteria, which are usually less than 5 um (Kubitschek et al. 1993), and 5-10 times larger than most human cells, which are 10-20 µm (Barrandon et al. 1985). Ciliates are more like a human cell than a bacterium because their DNA is encased inside an envelope-like membrane in a nucleus (actually in many nuclei, Lee et al. 2000) inside the cell, which bacteria do not have.

Ciliates generally eat other organisms, including algae, bacteria, and even other ciliates (Lee et al. 2000). They get the name “ciliate” from the hair-like cilia, structures that beat like tiny oars to propel the cell gracefully through water – even the small amounts of water between grains of soil! Ciliates live in all kinds of environments, from inside humans (yuck), to soils, to ephemeral rock pools on top of volcanoes, and even on glaciers (personal observation!).

An example of a ciliate (fig. 1) you may have seen in biology class is Paramecium multinucleatum. Why, you might wonder, is a lab focused on uncultured wild microbes in alpine soils writing about a Paramecium? Well, Paramecium putrinum, a cousin of sorts of the common “lab rat” P. multinucleatum, turned up in Antarctic cryoconite holes! Cryoconite holes are mud puddles that melt into glaciers. You can learn more about our research in Antarctic cryoconite holes at cryoholes.wordpress.com.

Mieczan and colleagues (2013) found P. putrinum in 30-40% of the cryoconite holes on a coastal Antarctic glacier, both in the sediment at the bottom of the hole and at the top. Having watched many a “lab rat” Paramecium cruising around a petri dish, their wide distribution inside the cryoconite holes does not surprise me. Having tried freezing and resuscitating my cultures of Paramecium multinucleatum, however, the ability of this species to live in Antarctica is surprising! Cryoconite holes there definitely freeze solid during the long, dark winter, so P. putrinum must have freeze-tolerant abilities that its lab rat cousin lacks.We find a few DNA sequences that map most closely to Paramecium in cryoconite holes in a dry valley of Antarctica (fig. 2-3), on the far side of the continent from where Mieczan and colleagues sampled. Although ciliates in general are some of our most abundant DNA there, Paramecium is not the most common – but it might be a similar Paramecium to the one Mieczan found!